Magnetofection is a transfection method that uses magnetic fields to concentrate particles containing vectors to target cells in the body. [1] Magnetofection has been adapted to a variety of vectors, including nucleic acids, non-viral transfection systems, and viruses. This method offers advantages such as high transfection efficiency and biocompatibility which are balanced with limitations.
The term magnetofection, currently trademarked by the company OZ Biosciences, combines the words magnetic and transfection. [2] Magnetofection uses nucleic acids associated with magnetic nanoparticles. These molecular complexes are then concentrated and transported into cells using an applied magnetic field.
The magnetic nanoparticles are typically made from iron oxide, which is fully biodegradable, using methods such as coprecipitation or microemulsion. [3] [4]
The nanoparticles are then combined with gene vectors (DNA, siRNA, ODN, virus, etc.). One method involves linking viral particles to magnetic particles using an avidin-biotin interaction. [5] Viruses can also bind to the nanoparticles via hydrophobic interaction. [6]
Another synthesis method involves coating magnetic nanoparticles with cationic lipids or polymers via salt-induced aggregation. For example, nanoparticles may be conjugated with the polyethylenimine (PEI), a positively charged polymer used commonly as a transfection agent. [7] The PEI solution must have a high pH during synthesis to encourage high gene expression. [8] The positively charged nanoparticles can then associate with negatively charged nucleic acids via electrostatic interaction. [9]
Magnetic particles loaded with vectors are concentrated on the target cells by the influence of an external magnetic field. The cells then take up genetic material naturally via endocytosis and pinocytosis. Consequently, membrane architecture and structure stays intact, in contrast to other physical transfection methods such as electroporation or gene guns that damage the cell membrane. [10]
The nucleic acids are then released into the cytoplasm by different mechanisms depending upon the formulation used:
Magnetofection works with cells that are not dividing or slowly dividing, meaning that the genetic materials can go to the cell nucleus without cell division. [11]
Magnetofection has been tested on a broad range of cell lines, hard-to-transfect and primary cells. [12] Several optimized and efficient magnetic nanoparticle formulations have been specifically developed for several types of applications such as DNA, siRNA, and primary neuron transfection as well as viral applications. [13]
Magnetofection research is currently in the preclinical stage. This technique has primarily been tested in vivo using plasmid DNA in mouse, rat, and rabbit models for applications in the hippocampus, subcutaneous tumors, lungs, spinal cord, and muscle. [14]
Some applications include:
Magnetofection attempts to unite the advantages of biochemical (cationic lipids or polymers) and physical (electroporation, gene gun) transfection methods. It allows for local delivery with high transfection efficiency, faster incubation time, and biocompatibility. [19]
Coupling magnetic nanoparticles to gene vectors results in hundreds-fold increase of the uptake of these vectors on a time scale of minutes, thus leading to high transfection efficiency. [20] Gene vector and magnetic nanoparticle complexes are transfected into cells after 10–15 minutes, which is faster than the 2–4 hours that other transfection methods require. [21] After 24, 48 or 72 hours, most of the particles are localized in the cytoplasm, in vacuoles (membranes surrounded structure into cells) and occasionally in the cell nucleus. [22]
Magnetic nanoparticles do not aggregate easily once the magnet is removed, and therefore are unlikely to block capillaries or cause thrombosis. [23] In addition, iron oxide is biodegradable, and the iron can be reused in hemoglobin or iron metabolism pathways. [24] [25]
Magnetic nanoparticle synthesis can sometimes lead to a wide range of differently sized particles. [26] The size of particles can influence their usefulness. Specifically, nanoparticles that are less than 10 nm or greater than 200 nm in size tend to be cleared from the body more quickly. [27]
While magnets can be used to localize magnetic nanoparticles to desired cells, this mechanism may be difficult to maintain in practice. The nanoparticles can be concentrated in 2D space such as on a culture plate or at the surface of the body, but it can be more difficult to localize them in the 3D space of the body. Magnetofection does not work well for organs or blood vessels far from the surface of the body, since the magnetic field weakens as distance increases. [28] [29] In addition, the user must consider the frequency and timing of applying the magnetic field, as the particles will not necessarily stay in the desired location once the magnet is removed. [30]
While iron oxide used to make nanoparticles is biodegradable, the toxicity of magnetic nanoparticles is still under investigation. Some research has found no signs of damage to cells, while others claim that small (< 2 nm) nanoparticles can diffuse across cell membranes and disrupt organelles. [31] [32]
In addition, very high concentrations of iron oxide can disrupt homeostasis and lead to iron overload, which can damage or alter DNA, affect cellular responses, and kill cells. [33] Lysosymes can also digest the nanoparticles and release free iron which can react with hydrogen peroxide to form free radicals, leading to cytotoxic, mutagenic, and carcinogenic effects. [34]
Nanomedicine is the medical application of nanotechnology. Nanomedicine ranges from the medical applications of nanomaterials and biological devices, to nanoelectronic biosensors, and even possible future applications of molecular nanotechnology such as biological machines. Current problems for nanomedicine involve understanding the issues related to toxicity and environmental impact of nanoscale materials.
Small interfering RNA (siRNA), sometimes known as short interfering RNA or silencing RNA, is a class of double-stranded non-coding RNA molecules, typically 20–24 base pairs in length, similar to microRNA (miRNA), and operating within the RNA interference (RNAi) pathway. It interferes with the expression of specific genes with complementary nucleotide sequences by degrading messenger RNA (mRNA) after transcription, preventing translation. It was discovered in 1998 by Andrew Fire at the Carnegie Institution for Science in Washington, D.C. and Craig Mello at the University of Massachusetts in Worcester.
Transfection is the process of deliberately introducing naked or purified nucleic acids into eukaryotic cells. It may also refer to other methods and cell types, although other terms are often preferred: "transformation" is typically used to describe non-viral DNA transfer in bacteria and non-animal eukaryotic cells, including plant cells. In animal cells, transfection is the preferred term as transformation is also used to refer to progression to a cancerous state (carcinogenesis) in these cells. Transduction is often used to describe virus-mediated gene transfer into prokaryotic cells.
Nanoid robotics, or for short, nanorobotics or nanobotics, is an emerging technology field creating machines or robots, which are called nanorobots or simply nanobots, whose components are at or near the scale of a nanometer. More specifically, nanorobotics refers to the nanotechnology engineering discipline of designing and building nanorobots with devices ranging in size from 0.1 to 10 micrometres and constructed of nanoscale or molecular components. The terms nanobot, nanoid, nanite, nanomachine and nanomite have also been used to describe such devices currently under research and development.
Cationic liposomes are spherical structures that contain positively charged lipids. Cationic liposomes can vary in size between 40 nm and 500 nm, and they can either have one lipid bilayer (monolamellar) or multiple lipid bilayers (multilamellar). The positive charge of the phospholipids allows cationic liposomes to form complexes with negatively charged nucleic acids through ionic interactions. Upon interacting with nucleic acids, cationic liposomes form clusters of aggregated vesicles. These interactions allow cationic liposomes to condense and encapsulate various therapeutic and diagnostic agents in their aqueous compartment or in their lipid bilayer. These cationic liposome-nucleic acid complexes are also referred to as lipoplexes. Due to the overall positive charge of cationic liposomes, they interact with negatively charged cell membranes more readily than classic liposomes. This positive charge can also create some issues in vivo, such as binding to plasma proteins in the bloodstream, which leads to opsonization. These issues can be reduced by optimizing the physical and chemical properties of cationic liposomes through their lipid composition. Cationic liposomes are increasingly being researched for use as delivery vectors in gene therapy due to their capability to efficiently transfect cells. A common application for cationic liposomes is cancer drug delivery.
Targeted drug delivery, sometimes called smart drug delivery, is a method of delivering medication to a patient in a manner that increases the concentration of the medication in some parts of the body relative to others. This means of delivery is largely founded on nanomedicine, which plans to employ nanoparticle-mediated drug delivery in order to combat the downfalls of conventional drug delivery. These nanoparticles would be loaded with drugs and targeted to specific parts of the body where there is solely diseased tissue, thereby avoiding interaction with healthy tissue. The goal of a targeted drug delivery system is to prolong, localize, target and have a protected drug interaction with the diseased tissue. The conventional drug delivery system is the absorption of the drug across a biological membrane, whereas the targeted release system releases the drug in a dosage form. The advantages to the targeted release system is the reduction in the frequency of the dosages taken by the patient, having a more uniform effect of the drug, reduction of drug side-effects, and reduced fluctuation in circulating drug levels. The disadvantage of the system is high cost, which makes productivity more difficult, and the reduced ability to adjust the dosages.
Gene delivery is the process of introducing foreign genetic material, such as DNA or RNA, into host cells. Gene delivery must reach the genome of the host cell to induce gene expression. Successful gene delivery requires the foreign gene delivery to remain stable within the host cell and can either integrate into the genome or replicate independently of it. This requires foreign DNA to be synthesized as part of a vector, which is designed to enter the desired host cell and deliver the transgene to that cell's genome. Vectors utilized as the method for gene delivery can be divided into two categories, recombinant viruses and synthetic vectors.
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Magnetic nanoparticles (MNPs) are a class of nanoparticle that can be manipulated using magnetic fields. Such particles commonly consist of two components, a magnetic material, often iron, nickel and cobalt, and a chemical component that has functionality. While nanoparticles are smaller than 1 micrometer in diameter, the larger microbeads are 0.5–500 micrometer in diameter. Magnetic nanoparticle clusters that are composed of a number of individual magnetic nanoparticles are known as magnetic nanobeads with a diameter of 50–200 nanometers. Magnetic nanoparticle clusters are a basis for their further magnetic assembly into magnetic nanochains. The magnetic nanoparticles have been the focus of much research recently because they possess attractive properties which could see potential use in catalysis including nanomaterial-based catalysts, biomedicine and tissue specific targeting, magnetically tunable colloidal photonic crystals, microfluidics, magnetic resonance imaging, magnetic particle imaging, data storage, environmental remediation, nanofluids, optical filters, defect sensor, magnetic cooling and cation sensors.
Magnet-assisted transfection is a transfection method which uses magnetic interactions to deliver DNA into target cells. Nucleic acids are associated with magnetic nanoparticles, and magnetic fields drive the nucleic acid-particle complexes into target cells, where the nucleic acids are released.
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Gene therapy utilizes the delivery of DNA into cells, which can be accomplished by several methods, summarized below. The two major classes of methods are those that use recombinant viruses and those that use naked DNA or DNA complexes.
Gene therapy is being studied for some forms of epilepsy. It relies on viral or non-viral vectors to deliver DNA or RNA to target brain areas where seizures arise, in order to prevent the development of epilepsy or to reduce the frequency and/or severity of seizures. Gene therapy has delivered promising results in early stage clinical trials for other neurological disorders such as Parkinson's disease, raising the hope that it will become a treatment for intractable epilepsy.
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Intracellular delivery is the process of introducing external materials into living cells. Materials that are delivered into cells include nucleic acids, proteins, peptides, impermeable small molecules, synthetic nanomaterials, organelles, and micron-scale tracers, devices and objects. Such molecules and materials can be used to investigate cellular behavior, engineer cell operations or correct a pathological function.
Hydrodynamic Delivery (HD) is a method of DNA insertion in rodent models. Genes are delivered via injection into the bloodstream of the animal, and are expressed in the liver. This protocol is helpful to determine gene function, regulate gene expression, and develop pharmaceuticals in vivo.
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